3D printing plant fish meat based on microscopic CT modeling and preparation method and application thereof

Abstract

The invention belongs to the technical field of food process application materials, and particularly relates to 3D printing plant fish meat based on microscopic CT modeling and a preparation method and application thereof. Real fish meat before 3D printing of artificial meat is subjected to digital modeling based on microscopic CT, meanwhile, vegetable protein paste is treated through heating alkali, protein molecules are unfolded and regathered, the solubility and water binding capacity of vegetable protein are enhanced, then the 3D printing performance of the vegetable protein is enhanced, and the pseudosciaena crocea simulated fish fillet with good texture characteristics is obtained.

Description

Technical Field

[0001] The present invention belongs to the technical field of food technology application materials, and specifically relates to a 3D printed plant fish meat based on micro-CT modeling, and a preparation method and application thereof. Background Art

[0002] As a high-quality protein source, aquatic products have a high feed conversion rate, low fat content, rich trace elements, and high protein content. They can meet future food needs and account for an increasing proportion of the diet. Artificial meat includes cell-cultured meat and plant-based meat. Cell-cultured meat is prepared by in vitro culture of stem cells and is a forward-looking study of edible meat, but it is expensive and difficult to market as soon as possible. Plant-based meat is made from raw materials such as soybeans, peas, and oats, and the molecular structure of plant protein is reconstructed into the fibrous structure of meat.

[0003] 3D printing technology is a manufacturing technology that converts digital models into physical objects. It builds three-dimensional objects by stacking materials layer by layer. Using 3D printing technology to shape artificial meat has become an effective solution to the poor consumer acceptance of artificial meat. Among them, improving the simulation degree of modeling before 3D printing is the top priority. Generally, 3D scanning is used for simulation 3D modeling. Three-dimensional scanning refers to a high-tech technology that integrates optical, mechanical, electrical and computer technologies. It is mainly used to scan the spatial shape, structure and color of objects to obtain the spatial coordinates of the surface of the object, and can convert the three-dimensional information of the object into a digital signal that can be directly processed by the computer. However, 3D scanning cannot simulate the internal structure, and it is not a feasible strategy for artificial meat with relatively high visual effects. Chinese patent CN114711326B provides a food printing material based on suspension 3D printing, its preparation method and application in artificial meat. After optimizing the material formula, the printing material and support material are retained at the same time, so that each of them plays a role in the entire structure, that is, the muscle structure and fat structure are simulated respectively. Chinese patent CN115039855B provides a method for 3D printing of beef mince for the elderly by using acoustic resonance to assist in processing beef mince. The beef mince is mixed with alkaline amino acids and marinated by acoustic resonance technology, and then 3D printed with different filling densities to obtain beef products that are more suitable for the elderly. The above existing technologies have improved the taste of artificial meat to a certain extent, but the internal structure is still quite different from real meat products and needs further improvement. Summary of the invention

[0004] The technical problem to be solved by the present invention is that general simulation 3D modeling mostly uses 3D scanning, but 3D scanning cannot simulate the internal structure and is not a feasible strategy for artificial meat with relatively high visual effects requirements.

[0005] In response to the problems of the prior art, the present invention provides a 3D printed plant fish meat based on micro-CT modeling, and a preparation method and application thereof. The real fish meat before 3D printing of artificial meat is digitally modeled based on micro-CT, and the plant protein paste is treated with heating alkali to cause the protein molecules to unfold and re-aggregate, thereby enhancing the solubility and water holding capacity of the plant protein, thereby enhancing its 3D printing performance, and obtaining a simulated yellow croaker fillet with good texture characteristics.

[0006] To achieve the above purpose, the technical solution of the present invention is: a preparation method of 3D printed plant fish meat based on micro-CT modeling, digital modeling of real fish meat before 3D printing of artificial meat based on micro-CT, and then 3D printing, 3D printing of artificial meat based on micro-CT scanning of real fish meat modeling helps artificial meat to obtain a more realistic fish meat shape to improve consumers' acceptance of the appearance of artificial meat. Among them, digital modeling includes the following steps:

[0007] (1) After pretreatment, fish samples were subjected to micro-CT scanning to obtain systematic slice images of the prepared samples;

[0008] (2) performing data processing and three-dimensional rendering on the micro-CT scanning data to obtain a corresponding 3D scanning simulation model after micro-CT data processing;

[0009] (3) After three-dimensional rendering, the surface mesh is extracted to obtain the three-dimensional data of large yellow croaker fillet;

[0010] (4) The surface of the large yellow croaker fillet three-dimensional data is smoothed to obtain the corresponding large yellow croaker fillet three-dimensional model after data processing.

[0011] Furthermore, step (1) fish sample pretreatment includes the following steps: cutting a 3-4 square centimeter fish fillet from the dorsal side of the fish; fixing with 4% PFA fixative at room temperature for 24 to 36 hours; then, washing the fillet with sterile saline; and staining with 3.75% Lugol's iodine solution for 7 to 14 days to enhance the contrast of the CT image. If the fish sample volume is too large and exceeds the volume range that can be accommodated and scanned by the micro-CT, it cannot be scanned and photographed; if the volume is too small, it cannot represent the general appearance of the fish; a short fixation time may result in uneven staining; a long staining time will result in a waste of resources; if the staining time is too short, the density of the fish's own soft tissue is too small, which will affect the CT imaging effect of capturing details; if the staining time is too long, the fish tissue will dehydrate and deform.

[0012] Furthermore, the micro-CT scanning parameters of step (1) are set as follows: (1) source voltage 80-120 kV, source current 180-220 μA; if the voltage and current are too high, it may cause overexposure, resulting in reduced image contrast and blurred details; if the voltage and current are too low, it may not be possible to penetrate the sample and obtain a complete internal structure image.

[0013] (2) Exposure time 450-550ms; Longer exposure time helps to improve the brightness and clarity of the image to a certain extent, especially for meat samples with complex structure and low contrast, which can make more details appear in the image. However, too long exposure time may also introduce more noise, affect image quality, and increase scanning time and radiation dose.

[0014] (3) Rotation step length 0.2-0.4°; a relatively small rotation step length means that a larger number of images are collected during the sample rotation process, and a more detailed tomographic image sequence can be obtained, thereby improving the accuracy and precision of three-dimensional reconstruction and better presenting the details of the internal microstructure of meat, such as the orientation of muscle fibers and the pores between tissues. However, a smaller rotation step length will increase the scanning time and data volume, and also place higher requirements on the performance and storage of the equipment.

[0015] (4) Image pixel size is 10-15 μm. Smaller pixel size requires more storage resources and processing time, but it can also provide higher spatial resolution, so that the reconstructed image can more clearly show the characteristics of the microstructure inside the meat, providing a more accurate model basis for subsequent 3D printing simulation.

[0016] Furthermore, in step (2), the micro-CT scan data is subjected to noise removal processing to obtain corresponding denoised micro-CT scan data; and then the result is converted into a binary format using interactive threshold segmentation.

[0017] Furthermore, the above threshold segmentation modifies the grayscale value extracted within the grayscale range (0 to 250). The extracted density will not affect the fat line used to identify the muscle. By limiting the grayscale range, the structure or tissue corresponding to the grayscale value within the range can be highlighted. And to a certain extent, the interference of other structures or backgrounds whose grayscale values ​​are not within this range can be suppressed, reducing the noise and clutter in the image, making the target structure more prominent, thereby better restoring its true structure in 3D reconstruction and improving the accuracy and reliability of the model.

[0018] Further, step (3) cuts the three-dimensional model of the large yellow croaker fillet after post-processing as a solid mesh. Select appropriate professional three-dimensional data processing and modeling software, and import the three-dimensional model of the large yellow croaker fillet obtained by post-processing into the selected software in a suitable format (such as STL, OBJ, etc.), to ensure that the model can be correctly displayed and operated in the software. Determine the position and direction of the cutting plane according to the needs of the actual application. Set cutting parameters such as cutting depth, cutting angle, cutting range, etc. according to requirements. The three-dimensional model of the large yellow croaker fillet can be flexibly adjusted according to the shape of the fillet and the purpose of the research. After confirming that the cutting plane and parameter settings are correct, perform the cutting operation. The software cuts the model according to the set parameters to generate the cut model part. Observe the cut model to check whether the cut surface is flat, whether the cutting position is accurate, and whether the expected cutting effect is achieved. If the cut surface is not smooth, there are excess fragments, etc., it is necessary to adjust and repair it. Perform necessary repair and optimization on the cut model, such as filling holes in the cut surface, smoothing the model surface, etc. Export the cut and optimized model in a suitable format for subsequent analysis, visualization or other applications.

[0019] Furthermore, the step of 3D printing plant fish meat includes the following steps: re-dissolving the denatured plant protein gel powder in water, then adding vegetable oil, and stirring evenly with an electric stirrer to obtain a plant protein-based 3D printing bio-ink; 3D printing the plant protein-based 3D printing bio-ink, and obtaining a gel texture-enhanced 3D printed plant fish meat after incubation;

[0020] The preparation of the denatured plant protein gel powder includes the following steps: dispersing the plant protein in water to obtain a plant protein paste, adjusting the pH value to 7-11 and heating it in a water bath for a certain period of time to form a denatured plant protein gel; freezing the plant protein paste for 24 hours and then vacuum-freezing and grinding to obtain a denatured plant protein gel powder. The present invention denatures the protein molecular structure by heating the plant protein paste with alkali, causing the protein to unfold and re-aggregate, thereby enhancing the solubility and water holding capacity of the plant protein, and thereby enhancing its 3D printing performance.

[0021] Furthermore, the above-mentioned plant protein is one or more of pea protein, yeast protein and soy protein.

[0022] Furthermore, the protein concentration in the vegetable protein paste is 14-22wt%, preferably 20wt%. The distance between the protein molecules in the paste obtained with a lower protein concentration is farther, and stronger molecular interactions are required to aggregate the proteins and significantly change the appearance of the paste, and the lubricating effect of a high content of water will reduce the gel strength of the protein aggregation system; when the protein concentration is higher, the distance between the protein molecules is closer and the density is higher, so more hydroxide ions need to be consumed to interact with the protein molecular chain, or cause a waste of resources.

[0023] Furthermore, the pH value was adjusted to 11.

[0024] Furthermore, the heating temperature is 80-100°C, preferably 80°C, and the heating time is selected from 10-30 minutes, preferably 25 minutes. Under low temperature or short heating time conditions, the protein cannot be fully unfolded to form a gel, and high temperature and long heating time will lead to chain reactions such as excessive denaturation of the protein and water loss.

[0025] Furthermore, when the denatured plant protein gel powder is redissolved in water, the proportion of the denatured plant protein gel powder is 19-22%, preferably 19.8%. Low solid content will affect the support of the product due to the fluidity and low viscosity brought by high water content, while high solid content will lead to excessive viscosity, thereby affecting the stability of the bio-ink when it is extruded into filaments by the nozzle.

[0026] Furthermore, the vegetable oil is selected from one or more of corn oil, peanut oil, soybean oil and linseed oil.

[0027] Furthermore, the proportion of vegetable oil in plant protein-based 3D printing bio-ink is 1-3%; in order to supplement nutrients and provide lubrication, too high a vegetable oil content will reduce the viscosity of the bio-ink and affect its supporting performance, while too low a content may cause extrusion jams.

[0028] Application of the above method in 3D printed products.

[0029] The plant protein-based 3D printing bio-ink prepared by the above method has excellent gel strength and 3D printing molding ability. It can be used to adapt to more complex 3D printing models to prepare 3D printed plant fish meat with good texture strength.

[0030] The plant protein-based 3D printed plant fish meat prepared by the above method is a gel texture-enhanced 3D printed plant fish meat product obtained by setting printing parameters and product shape and performing 3D printing.

[0031] Furthermore, the printing mode is set to concentric circle printing. In this mode, scanning model printing can minimize the impact of model fragments on the shape of the final product. The printing parameters are: print head diameter of 0.84mm, printing speed of 5-10mm / s, extrusion flow rate of 5-10mm 3 / s.

[0032] The beneficial effects of the present invention are:

[0033] (1) The present invention uses micro-CT to digitally model the real fish meat before 3D printing of artificial meat, and then performs 3D printing. The 3D printing of artificial meat based on the modeling of real fish meat scanned by micro-CT helps the artificial meat to obtain a more realistic fish meat shape to improve consumers' acceptance of the appearance of artificial meat. The traditional artificial meat production method is difficult to ensure the consistency of product quality, while the 3D printing technology based on micro-CT digital modeling can accurately print according to the preset digital model, ensuring that the structure and composition of each piece of artificial meat are highly consistent, thereby effectively improving the stability and consistency of product quality. Digital modeling provides a standardized digital template for the production of artificial meat. During the 3D printing process, it is only necessary to print according to the model parameters, which reduces the quality fluctuations caused by differences in manual operations in the traditional production process, improves the degree of automation of the production process, and helps to improve production efficiency and reduce production costs.

[0034] (2) The present invention treats the plant protein paste by heating alkali, causing the protein molecules to unfold and re-aggregate, thereby enhancing the solubility and water holding capacity of the plant protein, thereby enhancing its 3D printing performance, and obtaining a simulated large yellow croaker fillet with good texture characteristics. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 Particle size distribution results of denatured pea protein prepared at different pH values.

[0036] Figure 2 Dynamic shear scanning viscosity results of bio-ink prepared by denatured pea protein at different pH values.

[0037] Figure 3 Dynamic shear loss modulus results of bio-ink prepared by denatured pea protein at different pH values.

[0038] Figure 4 Printing effect diagram of bio-ink prepared by denatured pea protein with different pH values.

[0039] Figure 5 Scanning electron microscopy results of bio-ink prepared by denatured pea protein at different pH values.

[0040] Figure 6 Comparison of the texture results of 3D printed simulated yellow croaker fillets prepared with denatured pea protein at different pH values ​​and real yellow croaker fillets.

[0041] Figure 7 Schematic diagram of the model established by micro-CT scanning of real large yellow croaker fillets.

[0042] Figure 8 Schematic diagram of texture-enhanced 3D-printed simulated yellow croaker fillet after micro-CT scanning modeling. DETAILED DESCRIPTION

[0043] The following description is only a preferred embodiment of the present invention and is not intended to limit the scope of protection of the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

[0044] The following examples can be understood as a part of the local structure or method of the present invention, or can be understood as a combination of the examples to explain the connotation of the larger structure or method of the present invention. Unless otherwise specified, all raw materials of the present invention are purchased from commercial channels.

[0045] Embodiment 1:

[0046] A method for establishing a 3D printed plant fish model based on micro-CT comprises the following steps:

[0047] (1) After the fish meat samples were pretreated, micro-CT scanning was performed to obtain a systematic slice image of the prepared sample; wherein the fish meat sample pretreatment included the following steps: cutting a 3-4 cm2 fish fillet from the dorsal side of the fish; fixing it with 4% PFA fixative at room temperature for 24 hours; then, washing the fillet with sterile saline; and staining it with 3.75% Lugol's iodine solution for 7 days to enhance the contrast of the CT image. The micro-CT scanning parameters were set as follows: (1) source voltage 100 kV, source current 200 μA; (2) exposure time 500 ms; (3) rotation step 0.3°; (4) image pixel size 10.14 μm.

[0048] (2) De-noising the micro-CT scan data to obtain the corresponding de-noised micro-CT scan data; then the result is converted into a binary format using interactive threshold segmentation. The threshold segmentation modifies the grayscale value extracted from the grayscale range (138-235). The extracted density does not affect the fat line used to identify the muscle.

[0049] (3) Cut the three-dimensional model of large yellow croaker fillet after post-processing as a solid mesh. Select appropriate professional three-dimensional data processing and modeling software, and import the three-dimensional model of large yellow croaker fillet obtained by post-processing into the selected software in a suitable format (such as STL, OBJ, etc.) to ensure that the model can be correctly displayed and operated in the software. Determine the position and direction of the cutting plane according to the needs of the actual application. Set the cutting parameters according to the requirements, such as cutting depth, cutting angle, cutting range, etc. The three-dimensional model of large yellow croaker fillet can be flexibly adjusted according to the shape of the fillet and the purpose of the research. After confirming that the cutting plane and parameter settings are correct, perform the cutting operation. The software cuts the model according to the set parameters to generate the cut model part. Observe the cut model to check whether the cut surface is flat, whether the cutting position is accurate, and whether the expected cutting effect is achieved. If the cut surface is not smooth or there are excess fragments, etc., it is necessary to adjust and repair it. Perform necessary repairs and optimizations on the cut model, such as filling holes in the cut surface and smoothing the model surface. Export the cut and optimized model in a suitable format for subsequent analysis, visualization or other applications. The resulting model is as follows: Figure 7 shown.

[0050] Embodiment 2:

[0051] A method for preparing 3D printed plant fish meat based on pea protein structure enhancement, comprising the following steps:

[0052] The preparation of the denatured plant protein gel powder includes the following steps: dispersing 20g of pea protein in 80mL of distilled water, stirring with an electric stirrer at 2000r / min for 20min to fully hydrate the pea protein to form a pea protein paste; adjusting the pH to 7-11 during the stirring process, heating at 80°C in a water bath for 25min, and then refrigerating at -4°C for 18h to form a denatured plant protein gel; freezing the denatured plant protein gel in a -80°C refrigerator for 24h and then vacuum freeze-drying at -60°C, and obtaining a denatured plant protein gel powder after sufficient grinding; re-dissolving 19.8g of the denatured plant protein gel powder in 77.7mL of water, then adding 2.5mL of linseed oil, and stirring with an electric stirrer at 2000r / min for 30min to obtain a plant protein-based 3D printing bio-ink; 3D printing the plant protein-based 3D printing bio-ink to obtain a gel texture-enhanced 3D printed plant fish meat.

[0053] 3D printing plant fish meat: the denatured plant protein gel powder is re-dissolved in water, and then vegetable oil is added, and the mixture is stirred evenly with an electric stirrer to obtain a plant protein-based 3D printing bio-ink; the plant protein-based 3D printing bio-ink is 3D printed, and after incubation, a gel-textured 3D printed plant fish meat is obtained;

[0054] Among them, when the denatured plant protein gel powder is redissolved in water, the proportion of the denatured plant protein gel powder is 19.8%. The plant oil is selected from one or more of corn oil, peanut oil, soybean oil, and linseed oil. The proportion of plant oil in the plant protein-based 3D printing bio-ink is 2.5%. The plant protein-based 3D printing bio-ink obtains a gel texture to strengthen 3D printed plant fish meat.

[0055] The printing model used is set to a 3×3×3cm 3 The printing parameters are: the print head diameter is 0.84mm, the printing speed is 5-10mm / s, and the extrusion flow rate is 5-10mm 3 / s. The printing effect of the bio-ink prepared by pH 7-11 and undenatured pea protein gel is shown in Figure 2. Figure 4 shown.

[0056] Embodiment 3:

[0057] A method for preparing 3D printed plant fish meat based on micro-CT modeling, comprising the following steps:

[0058] (1) After the fish sample was pretreated, micro-CT scanning was performed to obtain a systematic slice image of the prepared sample; wherein the fish sample pretreatment included the following steps: cutting a 3-4 square centimeter fish fillet from the dorsal side of the fish; fixing it with 4% PFA fixative at room temperature for 24 hours; then, washing the fish fillet with sterile saline; and staining it with 3.75% Lugol's iodine solution for 7 days to enhance the contrast of the CT image. The micro-CT scanning parameters were set as follows: (1) source voltage 100 kV, source current 200 μA;

[0059] (2) Exposure time 500ms; (3) Rotation step 0.3°; (4) Image pixel size 10.14μm.

[0060] (2) De-noising the micro-CT scan data to obtain the corresponding de-noised micro-CT scan data; then the result is converted into a binary format using interactive threshold segmentation. The threshold segmentation modifies the grayscale value extracted from the grayscale range (141-229). The extracted density does not affect the fat line used to identify the muscle.

[0061] (3) After three-dimensional rendering, the processed three-dimensional model of the large yellow croaker fillet is cut as a solid mesh, and the surface mesh is extracted to obtain the three-dimensional data of the large yellow croaker fillet;

[0062] (4) The surface of the large yellow croaker fillet three-dimensional data is smoothed to obtain the corresponding large yellow croaker fillet three-dimensional model after data processing.

[0063] (5) 3D printing plant fish meat: the denatured plant protein gel powder is redissolved in water, and then vegetable oil and glutamine transaminase are added, and the mixture is stirred evenly with an electric stirrer to obtain a plant protein-based 3D printing bio-ink; the plant protein-based 3D printing bio-ink is 3D printed, and after incubation, a gel-textured 3D printed plant fish meat is obtained;

[0064] The preparation of the denatured vegetable protein gel powder includes the following steps: dispersing the vegetable protein in water to obtain a vegetable protein paste, adjusting the pH and heating it in a water bath for a certain period of time to form a denatured vegetable protein gel; freezing the vegetable protein paste for 24 hours and then freeze-drying it in a vacuum, and grinding it to obtain a denatured vegetable protein gel powder. The vegetable protein is one or more of pea protein, yeast protein and soy protein. The protein concentration in the vegetable protein paste is 20wt%. The pH value is adjusted to 11. The heating temperature is 80; the heating time is selected from 10 to 30 minutes; preferably 25 minutes.

[0065] Among them, when the denatured plant protein gel powder is redissolved in water, the proportion of the denatured plant protein gel powder is 19.8%. The plant oil is selected from one or more of corn oil, peanut oil, soybean oil, and linseed oil. The proportion of plant oil in the plant protein-based 3D printing bio-ink is 2.5%. After the plant protein-based 3D printing bio-ink is obtained, the gel texture is strengthened to print plant fish meat.

[0066] The printing mode is set to concentric circle printing. In this mode, scanning model printing can minimize the impact of model fragments on the shape of the final product. The printing parameters are: print head diameter is 0.84mm, printing speed is 10mm / s, and extrusion flow rate is 5-10mm 3 / s. The simulated yellow croaker fillet obtained by 3D printing based on micro-CT scanning modeling is as follows Figure 8 shown.

[0067] Comparative Example:

[0068] 3D printed plant-based fish meat without micro-CT scanning and texture enhancement.

[0069] Without texture-enhanced pea protein, the prepared bio-ink can be molded into products due to the protein's own support properties and high protein concentration, but a large number of bubbles will be involved due to high-speed stirring ( Figure 4 ), this product lacks the elasticity and adhesion that fish simulated texture should have ( Figure 6 ) and lack of attractiveness in appearance.

[0070] If micro-CT scans were not available, manually constructed models were used ( Figure 4), this model does not have the realism of scanning modeling, and is indistinguishable from the commonly processed products on the market. It has heavy traces of human processing. In order to attract consumers' attention, it requires additional investment of high manpower to design a 3D printed model, and there is no way to compare with real fish meat.

[0071] Verification results:

[0072] (I) Particle size analysis of denatured pea protein gel

[0073] Depend on Figure 1 It can be seen that in the native protein system, the most common peak is in the range of 100-1000nm, which is mainly due to the influence of electrostatic solid repulsion caused by surface charge (-32.17mV). When the pH value is adjusted to 7, the proportion of particles with a diameter of 100-1000nm in the dispersion decreases with the increase of pH value, and the highest density moves to the left. At pH 8, a peak appeared in the small particle area with a diameter of 10-100nm. The proportion of this peak increased at pH 9 and suddenly stopped at pH 10; afterwards, a small peak appeared at 1000-10000nm. Further increasing the pH value will cause the peak width of 100-1000nm to decrease, while the peak intensity of 1000-10000nm will increase.

[0074] (II) 3D printing bio-ink prepared from denatured pea protein

[0075] 1. Dynamic shear sweep viscosity

[0076] To analyze the printing stability, the rheological properties of the bio-ink were further measured. Viscosity results ( Figure 2 ) showed that the viscosity of all bio-inks decreased with increasing shear rate. This indicates that all of these bio-inks belong to pseudoplastic fluid systems with shear-thinning behavior, which meets the basic rheological properties required for 3D printing materials. At a given frequency (1 rad / s), the bio-ink prepared with native protein has the highest apparent viscosity (6.7×10 5 mPa-s), followed by pH 11 (6.3×10 5 mPa-s). Meanwhile, the bio-ink with poor printing performance has a significantly lower apparent viscosity at 1 rad / s.

[0077] 2. Dynamic shear rheological properties

[0078] Dynamic viscoelastic behavior results ( Figure 3), the storage modulus at pH 11 was the highest (1 rad / s), with high gel strength and weak fluidity. In the early stage of pH adjustment (pH 7, 8, and 9), the increase in pH did not significantly improve elasticity (G'). In addition, when the G' of these bio-inks was fitted to a power law model, their a values ​​spanned from 1000 to 5000 (Pa·rad -b s b )(Table), while their b values ​​(frequency dependence) remained below 0.16 (Table 1). The G'a value of the bio-ink containing pH 11 was the highest, followed by native protein, and the G'a value of the bio-ink at pH 7 was the lowest (p<0.05). The pH 7 sample with the lowest a value had printing results of failure to extrude or poor extrusion ( Figure 4 ). A higher a value indicates a stronger solid behavior. Such a high a value indicates that the bio-ink hardly flows and relies mostly on the pressure given by the plunger when it moves during the extrusion process. This will also make the extrusion process more controllable because the bio-ink is hardly extruded when the plunger is not moving. Overall, these myoseptum mimics exhibit strong viscoelastic solid behavior. Among them, the bio-ink obtained under pH 11 conditions has the greatest mechanical strength, which is reflected in its largest a value and G' over the entire frequency range.

[0079] Table 1

[0080]

[0081] (III) 3D printed plant fish meat with enhanced gel texture

[0082] 1. Cryo-SEM

[0083] The figure shows the SEM images of the bio-ink obtained by alkali treatment with different pH values ​​before and after enzyme treatment. The microstructure of the obtained gel is different when heated under different pH conditions. When heated, the pea protein begins to aggregate ( Figure 5 ); as the pH increases to pH 8, filamentous structures begin to form ( Figure 5 ), the protein gel structure was porous, indicating that the change of pH value can help the protein unfold under the assistance of heat. The gel at pH 11 showed a porous, thick-walled, densely cross-linked structure.

[0084] The SEM results show obvious changes in the samples before and after enzyme cross-linking. There is almost no change between the native protein and the protein paste prepared at pH 7, including the four samples before and after enzyme cross-linking. This seems to indicate that in the absence of an adequate environmental pH value, heating cannot cause sufficient active sites of the native protein to leak out, but rather the protein structure is folded inside the protein. At pH 9, more active sites are exposed, and a layered structure can be seen. Thanks to this exposure and unfolding effect, the pores after enzyme cross-linking at pH 11 are more ordered and smaller, and the connecting walls are thinner than those before enzyme cross-linking. The cross-linking effect of the enzyme strengthens the strength of the connecting wall, which makes the pores of the gel appear to be arranged in an orderly manner ( Figure 5 ).

[0085] 2. Texture

[0086] The texture parameters (hardness, adhesion, stickiness, elasticity, cohesion and chewiness) of simulated yellow croaker fillets and real yellow croaker fillets treated with various pH values ​​after enzyme cross-linking were measured. It can be seen that the increase in pH value and enzyme cross-linking have a positive effect on the texture characteristics of simulated yellow croaker fillets.

[0087] Before pH 11, the water retention capacity of protein gels, including native protein pastes, was very weak, which caused the protein gels to easily lose water under the action of enzyme incubation. Although the native protein sample and the pH 7 sample had higher hardness values ​​than the pH 11 sample, the elasticity of both was much lower than that of the pH 11 sample. This shows that the water lost during the long enzyme treatment caused the shell hardening phenomenon to increase the hardness, rather than the gel strength of the gel itself. The pH 8, 9, and 10 samples showed high adhesion and low hardness, elasticity, stickiness, and chewiness. These samples did not complete the paste-solid transformation within the active enzyme time. This may be because the protein was not fully unfolded during the heating process, and the protein chain was not unfolded, resulting in insufficient contact and function of the enzyme. Due to the excellent water retention capacity under pH 11 conditions, the high-strength gel network can strengthen the connection wall obtained during the heating alkali treatment under the action of long-term enzyme cross-linking, and lock the water with a dense porous structure. Therefore, the protein gel under pH 11 maintains its gel strength after enzyme cross-linking treatment, and the hardness, elasticity, cohesion, stickiness, and chewiness are all maintained at a high strength. It was found that the texture of the large yellow croaker fillet simulation prepared at pH 11 was the smallest difference from that of the real large yellow croaker fillet. In summary, only the large yellow croaker simulated fillet prepared under pH 11 conditions can maintain and strengthen its gel structure after enzyme cross-linking, effectively completing the gel-solid transformation to obtain large yellow croaker simulated fillet with good texture characteristics.

Claims

1. A method for preparing 3D printed plant fish meat based on micro-CT modeling, characterized in that Based on micro-CT, the real fish meat before 3D printing of artificial meat is digitally modeled and then 3D printed, wherein the digital modeling includes the following steps: (1) After pretreatment, fish samples were subjected to micro-CT scanning to obtain systematic slice images of the prepared samples; (2) performing data processing and three-dimensional rendering on the micro-CT scanning data to obtain a corresponding 3D scanning simulation model after micro-CT data processing; (3) After three-dimensional rendering, the surface mesh is extracted to obtain the three-dimensional data of large yellow croaker fillet; (4) The surface of the large yellow croaker fillet three-dimensional data is smoothed to obtain the corresponding large yellow croaker fillet three-dimensional model after data processing.

2. The preparation method according to claim 1, characterized in that: Step (1) fish sample pretreatment includes the following steps: cutting off 3-4 square centimeters of fish fillets from the dorsal side of the fish; fixing with 4% PFA fixative at room temperature for 24-36 hours; then, washing the fish fillets with sterile saline; and staining with 3.75% Lugol's iodine solution for 7-14 days to enhance the contrast of CT images; micro-CT scanning parameters are set as follows: (1) source voltage 80-120 kV, source current 180-220 μA; (2) exposure time 450-550 ms; (3) rotation step 0.2-0.4°; (4) image pixel size 10-15 μm.

3. The preparation method according to claim 1, characterized in that: Step (2) performs noise removal processing on the micro-CT scan data to obtain the corresponding denoised micro-CT scan data; then uses interactive threshold segmentation to convert the result into a binary format; threshold segmentation modifies the grayscale value extracted from the grayscale range (0 to 250).

4. The preparation method according to claim 1, characterized in that: Step (3) cuts the three-dimensional model of the large yellow croaker fillet after post-processing as a solid mesh.

5. The preparation method according to claim 1, characterized in that The steps of 3D printing plant fish meat include the following steps: re-dissolving the denatured plant protein gel powder in water, then adding vegetable oil, and stirring evenly with an electric stirrer to obtain plant protein-based 3D printing bio-ink; 3D printing the plant protein-based 3D printing bio-ink, and obtaining gel texture-enhanced 3D printed plant fish meat after incubation; The preparation of the denatured plant protein gel powder includes the following steps: dispersing the plant protein in water to obtain a plant protein paste, adjusting the pH value to 7-11 and heating it in a water bath for a certain period of time to form a denatured plant protein gel; freezing the plant protein paste for 24 hours and then freeze-drying it in a vacuum, and grinding it to obtain the denatured plant protein gel powder.

6. The preparation method according to claim 5, characterized in that: The vegetable protein is one or more of pea protein, yeast protein and soybean protein; the protein concentration in the vegetable protein paste is 14-22wt%.

7. The preparation method according to claim 5, characterized in that: The heating temperature is 80-100° C.; the heating time is selected from 10 to 30 minutes; and the plant protein-based 3D printing bio-ink is incubated at 55° C. for 10 hours to obtain a gel texture-reinforced 3D printed plant fish meat.

8. The preparation method according to claim 5, characterized in that: When the denatured plant protein gel powder is redissolved in water, the proportion of the denatured plant protein gel powder is 19-22%; the proportion of vegetable oil in the plant protein-based 3D printing biological ink is 1-3%.

9. Application of the method according to any one of claims 1 to 8 in 3D printed products.

10. A plant protein-based 3D printing bio-ink prepared by any method of claims 5-8.

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